Throughout this application, various references are cited in parentheses to describe more fully the state of the art to which this invention pertains. The disclosure of these references are hereby incorporated by reference into the present disclosure.
Rheumatoid arthritis (RA) is a prevalent autoimmune disease characterized by synovial inflammation and pannus formation which can lead to cartilage and bone degradation. This debilitating condition affects nearly 1% of the population. Genetic susceptibility to this disease is associated with MHC class II molecules that contain an amino acid motif known as the shared epitope (SE) and are designated as HLA-DR molecules (Gregersen, P. K., J. Silver, R. J. Winchester. 1987. The shared epitope hypothesis. An approach to understanding the molecular genetics of susceptibility to rheumatoid arthritis. Arthritis Rheum. 30:1205, Zhou, Z., H. A. Menard. 2002. Autoantigenic posttranslational modifications of proteins: does it apply to rheumatoid arthritis? Curr. Opin. Rheumatol. 14:250). The shared epitope, expressed by the amino acid residues Q/R, K/R, R, A, A, is positively charged and forms one of the major peptide anchoring pockets (known as P4) of the MHC class II molecules.
Previous reports have suggested that a distinct feature of a putative pathogenic peptide involved in RA may be the presence of the negatively charged side-chain at P4 (interacting with the shared epitope) (Hammer J., Gallazzi F., Bono E., Karr R. W., Guenot J., Valsasnini P., Nagy Z. A., Sinigaglia F. 1995. Peptide binding specificity of HLA-DR4 molecules: correlation with rheumatoid arthritis association. J. Exp. Med. 181:1847). This is based on the fact that certain MHC class II molecules, HLA-DR*0401 and HLA-DR*0404 (allelic variants) have a substantially higher affinity for aspartic and glutamic acid amino acid residues at the P4 pocket than the RA non-associated HLA-DR*0402 molecule. However, after analysis of multiple DRB1 pocket profiles it can be found that some RA non-associated alleles have a higher affinity for negatively charged amino acids at their P4 pockets than even HLA-DR *0101, HLA-DR *0401, and HLA-DR *0404, such as HLA-DR *0301 (Sturniolo T, Bono E, Ding J, Raddrizzani L, Tuereci O, Sahin U, Braxenthaler M, Gallazzi F, Protti M P, Sinigaglia F, Hammer J. 1999. Generation of tissue-specific and promiscuous HLA ligand databases using DNA microarrays and virtual HLA class II matrices. Nat. Biotechnol. 17:555). Some MHC molecules appear to be protective against disease (e.g. HLA-DR *0402), rather than simply non-associated, suggesting that a passive role for these alleles in peptide binding may not occur (Reviron, D., A. Perdriger, E. Toussirot, D. Wendling, N. Balandraud, S. Guis, G. Semana, P. Tiberghien, P. Mercier, J. Roudier. 2001. Influence of shared epitope-negative HLA-DRB1 alleles on genetic susceptibility to rheumatoid arthritis. Arthritis Rheum. 44:535). Instead, protective alleles (MHC molecules) may bind a putative pathogenic peptide with high enough affinity as to induce negative T cell selection, or to establish peripheral tolerance.
The MHC class II molecules with the shared epitope may participate in disease pathogenesis by selectively binding arthritogenic peptides for presentation to autoreactive CD4+ T cells. Currently, the nature of the autoantigen responsible for RA is not known. While many candidate autoantigens have been investigated in the context of RA associated MHC, a common disease specific target of the CD4+ T cell and B cell immune response remains elusive. Recent studies have shown that RA patients have a subset of IgG autoantibodies that are both sensitive and specific (>90%) for the diagnosis of RA. The target of these autoantibodies is citrulline, a post-translationally modified arginine (deiminated arginine) found within the context of certain protein/peptide sequences (van Venrooij, W. J., G. J. Pruijn. 2000. Citrullination: a small change for a protein with great consequences for rheumatoid arthritis. Arthritis Res. 2:249). Citrulline is the essential antigenic epitope target of anti-perinuclear, anti-keratin, anti-filaggrin, anti-cyclic citrullinated peptide, and anti-Sa antibodies (van Venrooij, W. J., G. J. Pruijn. 2000. Citrullination: a small change for a protein with great consequences for rheumatoid arthritis. Arthritis Res. 2:249; Zhou, Z., H. A. Menard. 2002. Autoantigenic posttranslational modifications of proteins: does it apply to rheumatoid arthritis? Curr. Opin. Rheumatol. 14:250). These antibodies target citrulline within a number of different proteins, the joint derived targets appears to be vimentin and fibrin(ogen) (Menard, H. A., E. Lapointe, M. D. Rochdi, Z. J. Zhou. 2000. Insights into rheumatoid arthritis derived from the Sa immune system. Arthritis Res. 2:429; Christine Masson-Bessiere et al., 2001. The Major Synovial Targets of the Rheumatoid arthritis-Specific Antifilaggrin Autoantibodies Are Deiminated Forms of the α and β Chains of Fibrin 1 The Journal of Immunology 166: 4177-4184). It is also observed that anti-citrulline antibody production is significantly associated with the presence of the MHC shared epitope in R A patients (Goldbach-Mansky R, Lee J, McCoy A, Hoxworth J, Yarboro C, Smolen J S, Steiner G, Rosen A, Zhang C, Menard H A, Zhou Z J, Palosuo T, Van Venrooij W J, Wilder R L, Klippel J H, Schumacher H R Jr, El-Gabalawy H S. 2000. Rheumatoid arthritis associated autoantibodies in patients with synovitis of recent onset. Arthritis Res. 2:236; Bas S, Perneger T V, Mikhnevitch E, Seitz M, Tiercy J M, Roux-Lombard P, Guerne P A. 2000. Association of rheumatoid factors and anti-filaggrin antibodies with severity of erosions in rheumatoid arthritis. Rheumatology (Oxford). 39: 1082).
The Applicant has now demonstrated that a unique interaction exists between the shared epitope of MHC class II molecules and the amino acid citrulline. This interaction is involved in generating T cell responses and subsequently B cell responses to these citrullinated antigens in autoimmune diseases where a patient expresses the shared epitope. Furthermore, the Applicant has now identified novel citrullinated antigens that evoke a T cell response leading to inflammation and the development of autoimmune disease such as rheumatoid arthritis as well as multiple sclerosis.